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Figure 1. C18 RP-HPLC purification of C. longurionis, showing peptides Lo6/7a and Lo6/b. The brown line shows the acetonitrile gradient.
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Figure 2. Purification of crude venom of C. asiaticus. (A) RP-HPLC chromatogram of C. asiaticus venom. The peak indicated in the box was collected for further purification; (B) Ion exchange chromatogram of the peak fractions collected in the first purification step. The indicated peaks were subjected to another RP-HPLC purification step; (C) RP-HPLC1 chromatogram as indicated in (B). Edman degradation of the first peak, Asi14a, revealed the amino acid sequence as described; (D) RP-HPLC2 chromatogram, as indicated in Figure 2B. Edman degradation of this peak, Asi3a, revealed the amino acid sequence as described above. The brown lines in (A,C,D) are acetonitrile gradients. The line in (B) shows the ion exchange gradient.
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Figure 3. RP-HPLC purification of AusB (gel filtration fraction). The brown line shows the acetonitrile gradient.
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Figure 4. Electrophysiological screening of Lo6/7a and Lo6/7b. * Represents traces after toxin application (1 μM) and overlapping control traces before toxin application.
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Figure 5. Electrophysiological screening of Asi3a on NaV and KV channels (10 μM) and nAChRs (5 μM). * Represents traces after toxin application and overlapping control traces before toxin application.
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Figure 6. Electrophysiological screening of Asi14a on KV channels (10 μM) and nAChRs (5 μM). * Represents traces after toxin application and overlapping control traces before toxin application.
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Figure 7. Electrophysiological screening of AusB (10 μM) on a panel of NaV, KV and nAChRs. * Represents traces after toxin application and overlapping control traces before toxin application.
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Figure 8. Alignment of Lo6/7a with conotoxins, sorted by the percentage of identity (indicated in the left column). References: Pr6c (C. parius, [22]), Vc7.4 (C. victoriae, [21]), a convulsant peptide from C. textile [20,23], Tx6.5 (C. textile, [24]), Vc7.3 (C. victoriae, [21]) and Cl6a (C. californicus, [25]).
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Figure 9. Alignment of Lo6/7b with conotoxins, sorted by percentage of identify (indicated in the left column). References: E6.2 (C. ermineus, unknown ref.); LtVIC (C. litteratus, [26]); Vc6.28 (C. victoriae, [21]); M1 (C. militaris, [27]); Conotoxin-3 (C. striatus, [28]); Bu9 (C. bullatus, [29]); Pu6.5 (C. pulicarius, [30]); Pu6.6 (C. pulicarius, [30]).
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Figure 10. Alignment of Asi3a with conotoxins, sorted by percentage of identity (indicated in the left column). References: Pr3a (C. parius, [22]), ArMMSK-01 (C. arenatus, [31]); Pu3.7 (C. pulicarius, [30]); Pu3.3 (C. pulicarius, [24]); Pu3.4 (C. pulicarius, [32]); BetTXIb (C. betulinus, [33]); TsMMSK-021 (C. tessulatus, [31]).
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